Project Summary: One major obstacle in treating spinal cord injury (SCI) is to replenish neurons that are lost during the course of injury and to restore local neuronal circuitry for functional repair. Stem cell transplantation therapy once showed great promise. However, its efficacy has not been satisfactory in clinical trials; immunorejection and ethical issues remain problematic. In vivo reprogramming is emerging as a potentially new breakthrough in regenerative medicine. This innovative technology literally converts endogenous glial cells into functional neurons for repair purposes, bypassing the challenging questions that stem cell replacement therapies are facing. Previous research from the PI?s lab has demonstrated that reactive astrocytes can be successfully converted in situ into functional neurons in both injured brain and brain of a model for Alzheimer?s disease by overexpression of a single transcription factor NeuroD1 (Guo et al, 2014; BEST of 2014 Article in Cell Stem Cell). However, molecular mechanisms of the reprogramming process remain elusive. MicroRNAs (miRNAs) are small non-coding RNAs that play pivotal roles during neural development and diseases. The function of miRNAs could be extensive given that one miRNA may regulate many target genes through the unique imperfect base-pairing mechanism exerting a global impact on the gene expression profile in a cell. In this proposal, the PI will combine his expertise on SCI and miRNA, and examine miRNA function during NeuroD1-mediated neuronal conversion in a mouse model of SCI. The central hypothesis of this proposal is that miRNAs play essential roles during the neuronal reprogramming process, and that forced expression of miRNAs, as well as NeuroD1, can reprogram reactive astrocytes into neurons for functional repair after SCI. The PI proposes three specific aims: 1) To determine miRNA function during NeuroD1-mediated neuronal conversion in the injured spinal cord; 2) To determine conversion of reactive astrocytes into neurons by miRNAs after SCI; 3) To determine functional integration of miRNA-converted neurons and their effects on animal?s behavior after contusive SCI. Completion of this proposal will show feasibility of miRNA-mediated glia-to-neuron conversion and lay out foundation for therapeutic application of this small RNA molecule as a synthetic drug in the future. The PI believes that this proposal will lead to a novel therapeutic treatment for SCI as well as other neurological disorders.